The long-term goal of our research is to understand the molecular and biochemical mechanisms by which histamine exerts its effects on the mammalian heart. The present proposal is designed to test the hypothesis that histamine, acting through the H2 receptor in ventricular muscle, increases cAMP levels, activates cAMP-dependent protein kinase, and stimulates the phosphorylation of sarcolemma proteins and that these effects are correlated with the positive inotropic actions of this agent. There are two major goals of the proposal: (1) Correlation of the positive inotropic effect of H2 agonists with changes in the protein kinase activity ratio. We will employ a series of guinea pig and rabbit hearts from fetal to adult and will determine their maximal inotropic response to H2 agonists using the Langendorff procedure. This response will be correlated with measurements of protein kinase and phosphorylase activity in hearts freeze-clamped at the time of maximum inotropy. An attempt will be made to study separately the particulate and soluble protein kinases to see whether the former might be more closely related to the contractile changes than the latter. On the basis of our previous work, we believed that the use of animals at different ages will provide a gradation in mechanical and biochemical responses and will therefore provide support for or against a cause-effect relationship between protein kinase activation and inotropy. (2) Correlation of the positive inotropic effect of H2 agonists with the phosphorylation of sarcolemma proteins. We will develop procedures for preparing highly purified sarcolemma membranes from guinea pig and rabbit hearts and will determine the nature of the membrane proteins phosphorylated by cAMP-dependent protein kinase and their relationship to the membrane proteins phosphorylated in the intact perfused heart in response to agonists. Correlation of the magnitude of the inotropic effect and the degree of phosphorylation in the perfused heart will provide support for the role of sarcolemma protein phosporylation in the mechanism of the contractile response to histamine. The ultimate goal of our research is to identify the substrates phosphorylated and their role in sarcolemmal function.